Preparation of baked product from dough

ABSTRACT

Addition of an oxidase and a protein disulfide isomerase (PDI) to a dough has a syn-ergistic effect on the loaf volume of a baked product made from the dough.

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a 35 U.S.C. 371 national application ofPCT/DK01/00646 filed Oct. 4, 2001 (the international application waspublished under PCT Article 21 (2) in English), which claims priority orthe benefit under 35 U.S.C. 119 of Danish application no. PA 2000 01521filed Oct.12, 2000 and U.S. provisional application no. 60/241,139 filedOct.17,2000, the contents of which are fully incorporated herein byreference.

FIELD OF THE INVENTION

[0002] The present invention relates to a process for preparing a bakedproduct made from dough. More particularly, it relates to a process forpreparing a baked product with an increased loaf volume.

BACKGROUND OF THE INVENTION

[0003] In the preparation of bread and other baked products from dough,it is generally desirable to increase the volume of the baked product.

[0004] The prior art discloses the addition to dough of various oxidaseswhich act on a substrate in the dough to form hydrogen peroxide, e.g. toobtain a larger loaf volume. A well known example is glucose oxidasefrom Aspergillus niger which acts on glucose, maltose etc. in the dough.

[0005] WO 9500636 discloses the use of a protein disulfide isomerase forstrengthening of gluten in bakery or pastry products.

[0006] WO 9529996 and WO 9931990 disclose the addition to dough ofcertain oxidases in combination with a protein disulfide isomerase.

SUMMARY OF THE INVENTION

[0007] The inventors have found that the addition of an oxidase and aprotein disulfide isomerase (PDI) to a dough has a synergistic effect onthe loaf volume of a baked product made from the dough.

[0008] Accordingly, the invention provides a process of preparing abaked product by adding a hydrogen peroxide-forming oxidase and aprotein disulfide isomerase to a dough, and baking the dough. Theoxidase is a hydrogen peroxide-forming oxidase except the following:

[0009] a) a glucose oxidase having optimum activity at pH 6-7 and havingmore than 75% of maximum activity at pH 8, and

[0010] b) a carbohydrate oxidase which has a higher activity on anoligosaccharide having a degree of polymerization of 2 or higher as asubstrate than on the corresponding monosaccharide.

[0011] The invention also provides a baking composition for use in theprocess.

DETAILED DESCRIPTION OF THE INVENTION

[0012] Hydrogen Peroxide-forming Oxidase

[0013] The following are some examples of oxidases with EC numbers(according to Enzyme Nomenclature): A monosaccharide oxidase such asglucose oxidase (EC 1.1.3.4), hexose oxidase (EC 1.1.3.5), galactoseoxidase (EC 1.1.3.9) or pyranose oxidase (EC 1.1.3.10). A deaminatingoxidase such as L-amino acid oxidase (EC 1.4.3.2), amine oxidase (EC1.4.3.4).

[0014] The oxidase may particularly have optimum activity at pH 4-5.5.

[0015] The glucose oxidase may be derived from a strain of Aspergillusor Penicillium, particularly A. niger, P. notatum, P. amagasakiense orP. vitale. The hexose oxidase may be one described in EP 833563. Thepyranose oxidase may be one described in WO 9722257, e.g. derived fromTrametes, particularly T. hirsuta. The galactose oxidase may be onedescribed in WO 0050606.

[0016] The deaminating oxidase may be one described in WO 9721351, e.g.a benzylamine oxidase derived from Pichia, particularly P. pastoris.

[0017] The glucose oxidase will typically be added in an amountcorresponding to 20-1,000 GODU/kg flour, particularly 75-300 GODU/kg(GODU activity unit defined below). For other oxidases, a similar assaymay be used by substituting the substrate.

[0018] A substrate of the oxidase may already be present in the dough(e.g. glucose or maltose), or it may be added.

[0019] Assay for Glucose Oxidase Activity (GODU)

[0020] Glucose oxidase oxidizes glucose in the presence of oxygen toform gluconolactone and hydrogen peroxide. This hydrogen peroxideoxidizes ABTS-R (2,2′-azino-di-[ethylbenzthiazoline-6-sulphonate]) inthe presence of peroxidase. This generates a blue-green color which ismeasured using a photometer at 418 nm.

[0021] Reaction conditions are 90 mM glucose, 0.1 M acetate buffer, pH5.6, incubation temperature 30° C., and reaction time 34 seconds. 1 unit(GODU) is the amount of enzyme which produces 1 μmole hydrogen peroxideper minute at these conditions.

[0022] Protein Disulfide Isomerase (PDI)

[0023] The PDI may particularly have optimum activity at pH 4-5.5.

[0024] The PDI may be from a mammalian or microbial source, e.g. fromyeast or filamentous fungi, such as Aspergillus, Saccharomyces orSchizosaccharomyces, particularly A. oryzae, A. niger, Saccharomycescerevisiae or Schizosaccharomyces pombe.

[0025] The PDI will typically be added in an amount of 0.2-50,particularly 1-10 mg/kg flour (calculated as pure enzyme protein).

[0026] Dough

[0027] The dough generally comprises wheat meal or wheat flour and/orother types of meal, flour or starch such as corn flour, corn starch,rye meal, rye flour, oat flour, oat meal, soy flour, sorghum meal,sorghum flour, potato meal, potato flour or potato starch.

[0028] The dough may be fresh, frozen or par-baked.

[0029] The dough is typically leavened e.g. by adding chemical leaveningagents or yeast, usually Saccharomyces cerevisiae (baker's yeast).

[0030] The dough may be a laminated dough.

[0031] The dough may also comprise other conventional dough ingredients,e.g.: proteins, such as milk powder, gluten, and soy; eggs (either wholeeggs, egg yolks or egg whites); an oxidant such as ascorbic acid,potassium bromate, potassium iodate, azodicarbonamide (ADA) or ammoniumpersulfate; an amino acid such as L-cysteine; a sugar; a salt such assodium chloride, calcium acetate, sodium sulfate or calcium sulfate. Thedough may comprise fat (triglyceride) such as granulated fat orshortening. The dough may further comprise an emulsifier such as amonoglyceride.

[0032] Baked Product

[0033] The process of the invention may be used for any kind of bakedproduct prepared from dough, either of a soft or a crisp character,either of a white, light or dark type. Examples are bread (in particularwhite, whole-meal or rye bread), typically in the form of loaves orrolls, French baguette-type bread, pita bread, tortillas, cakes,pancakes, biscuits, cookies, pie crusts, crisp bread, steamed bread,pizza and the like.

[0034] Baking Composition

[0035] The baking composition comprises an oxidase, a PDI and optionallyan additional enzyme as described below.

[0036] The baking composition may be an enzyme preparation, e.g. in theform of a granulate or agglomerated powder. It may have a narrowparticle size distribution with more than 95% (by weight) of theparticles in the range from 25 to 500 μm. Granulates and agglomeratedpowders may be prepared by conventional methods, e.g. by spraying theamylase onto a carrier in a fluid-bed granulator. The carrier mayconsist of particulate cores having a suitable particle size. Thecarrier may be soluble or insoluble, e.g. a salt (such as NaCl or sodiumsulfate), a sugar (such as sucrose or lactose), a sugar alcohol (such assorbitol), starch, rice, corn grits, or soy.

[0037] The baking composition may, in addition to enzymes, compriseother baking ingredients, particularly flour. Thus, the composition maybe a dough or a flour pre-mix.

[0038] Additional Enzyme

[0039] Optionally, an additional enzyme may be used together with thePDI and the oxidase.

[0040] The additional enzyme may be an amylase, a cyclodextringlucanotransferase, a peptidase, in particular an exopeptidase, atransglutaminase, a lipase, a phospholipase, a cellulase, ahemicellulase, a protease, a glycosyltransferase, a branching enzyme(1,4-α-glucan branching enzyme) or a second oxidoreductase (in additionto the hydrogen peroxide-forming oxidase).

[0041] The additional enzyme may be of any origin, including mammalianand plant, and preferably of microbial (bacterial, yeast or fungal)origin.

[0042] The amylase may be fungal or bacterial, e.g. a maltogenicalpha-amylase from B. stearothermophilus or an alpha-amylase fromBacillus, e.g. B. lichenifornis or B. amyloliquefaciens, a beta-amylase,e.g. from plant (e.g. soy bean) or from microbial sources (e.g.Bacillus), a glucoamylase, e.g. from A. niger, or a fungalalpha-amylase, e.g. from A. oryzae.

[0043] The hemicellulase may be a pentosanase, e.g. a xylanase which maybe of microbial origin, e.g. derived from a bacterium or fungus, such asa strain of Aspergillus, in particular of A. aculeatus, A. niger, A.awamori, or A. tubigensis, from a strain of Trichoderma, e.g. T. reesei,or from a strain of Humicola, e.g. H. insolens.

[0044] The protease may be from Bacillus, e.g. B. amyloliquefaciens.

[0045] The lipase may be derived from a strain of Thermomyces(Humicola), Rhizomucor, Candida, Aspergillus, Rhizopus, or Pseudomonas,in particular from T. lanuginosus (H. ianuginosa), Rhizomucor miehei, C.antarctica, A. niger, Rhizopus delemar, Rhizopus arrhizus or P. cepacia.

[0046] The phospholipase may have phospholipase A1 or A2 orlysophospholipase activity; it may or may not have lipase activity. Itmay be of animal origin, e.g. from pancreas, snake venom or bee venom,or it may be of microbial origin, e.g. from filamentous fungi, yeast orbacteria, such as Aspergillus or Fusarium, e.g. A. niger, A. oryzae orF. oxysporum. Also, the variants described in WO 0032758 may be used.

[0047] The second oxidoreductase may be a peroxidase, a laccase or alipoxygenase.

EXAMPLES Example 1

[0048] Bread was made according to a micro scale baking system using 12g meneba flour per dough. Ascorbic acid (30 ppm), fungal alpha-amylase(Fungamyl at 10 FAU/kg flour), and xylanase (100 FXU/kg flour) wereadded to each dough.

[0049] Baking were made with and without addition of PDI and glucoseoxidase to the dough, as indicated below. The glucose oxidase was fromAspergillus niger, and the PDI was a variant derived from Aspergillus.The volume of each loaf was measured and is expressed as specificvolume. PDI dosage Glucose oxidase Specific mg enzyme dosage volumeprotein/kg flour GODU/kg flour ml/g Control 0 0 3.6 ± 0.2 2.5 0 3.3 ±0.1 Reference 5 0 3.8 ± 0.1 0 200 3.8 ± 0.2 Invention 2.5 200 4.0 ± 0.15 200 4.3 ± 0.1

[0050] The results show that the volume improvement obtained with thecombination of glucose oxidase and PDI compared to the control washigher than the sum of the volume improvement obtained with either ofthe enzymes alone.

1. A process of preparing a baked product comprising: a) adding to adough: i) a hydrogen peroxide-forming oxidase except the following: a) aglucose oxidase having optimum activity at pH 6-7 and having more than75% of maximum activity at pH 8, and b) a carbohydrate oxidase which hasa higher activity on an oligosaccharide having a degree ofpolymerization of 2 or higher as a substrate than on the correspondingmonosaccharide, ii) and a protein disulfide isomerase, and b) baking thedough.
 2. The process of the preceding claim wherein the oxidase hasoptimum activity at pH 4-5.5.
 3. The process of either preceding claimwherein the oxidase is a glucose oxidase, particularly one derived fromAspergillus niger.
 4. The process of any preceding claim wherein theprotein disulfide isomerase is derived from Aspergillus, particularly A.niger or A. oryzae.
 5. A baking composition comprising: a) a hydrogenperoxide-forming oxidase except the following: i) a glucose oxidasehaving optimum activity at pH 6-7 and having more than 75% of maximumactivity at pH 8, and ii) a carbohydrate oxidase which has a higheractivity on an oligosaccharide having a degree of polymerization of 2 orhigher as a substrate than on the corresponding monosaccharide, b) and aprotein disulfide isomerase.
 6. The baking composition of the precedingclaim wherein the oxidase has optimum activity at pH 4-5.5.
 7. Thebaking composition of claim 5 or 6 which further comprises flour.
 8. Thebaking composition of the preceding claim which is a dough, a flourcomposition, or a flour pre-mix.
 9. The baking composition of any ofclaims 5-8 wherein the oxidase is a glucose oxidase, particularly onederived from Aspergillus niger.
 10. The baking composition of any ofclaims 5-9 wherein the protein disulfide isomerase is derived fromAspergillus, particularly A. niger or A. oryzae.